Takao Kida

Transgenic tomato plant carrying a gene for NADP-dependent glutamate dehydrogenase (gdha) from Aspergillus nidulans

Abstract Tomato plants were transformed with gene constructs that contained the gdhA gene for NADP-dependent glutamate dehydrogenase from Aspergillus nidulans coupled in the sense orientation with the constitutively active 35S promoter from cauliflower mosaic virus. Four independent transformants, which had one or several copies of the gene in their genomes, were obtained. In these transgenic lines, high-level expression of gdhA mRNA was detected in leaves and fruits, and NADP–GDH activity was detected at high levels in leaves. In the tomatoes from 6 successive weeks after the first flowering, the levels of total free amino acids in transgenic fruits were higher (2- to 3-fold) than that in controls. In particular, the level of glutamate was about twice that in control fruits.

In vitro propagation utilizing suspension cultures of meristematic nodular cell clumps and chromosome stability of Lilium×formolongi hort.

Abstract Suspension cultures composed of meristematic nodular cell clumps of Lilium×formolongi hort. line R13 have been maintained in liquid MS medium containing 1 mg/l picloram for over 4 years. All of the cell clumps checked were diploid and these clumps maintained a high potential for shoot regeneration. The fresh weight of meristematic nodular cell clumps increased 2.5-fold during 14 days of cultivation in a flask or bioreactor. The sucrose concentration in the regeneration medium and the addition of BA affected shoot regeneration from the cell clumps. More than 85% of the clumps produced shoots on solidified regeneration medium (1/2 MS medium containing 5–10 g/l sucrose) after 2 months of culture. A sucrose concentration more than 20 g/l resulted in the reduction of shoot regeneration. Addition of BA to the solidified medium was effective in increasing shoot numbers and stimulating the induction of shoots. The plantlets regenerated from cell clumps were all diploid with 24 chromosomes and grew normally to flowering. It is expected that about 4.5×1010 plantlets per year could be produced from 1 g of meristematic nodular cell clumps through this propagation method.

Effect of sugar type on the efficiency of plant regeneration from protoplasts isolated from shoot tip-derived meristematic nodular cell clumps of Lilium x formolongi hort.

SummarySuspension cultures composed of meristematic nodular cell clumps of Lilium x formolongi hort were established from shoot tips placed on MS medium supplemented with 1 mg/l picloram and 30 g/l sucrose, glucose, fructose or sorbitol. Protoplasts isolated from these cultures were embedded in 1 g/l gellan gumsolidified 1/2MS medium with 1 mg/l picloram and the different kinds of sugars at 0.5 M, and cultured at 25 °C in the dark. The highest plating efficiency (13.7%) was obtained when the protoplasts were isolated from the cell clumps which had been subcultured in MS medium containing glucose and were likewise cultured in MS medium supplemented with 0.5 M glucose. Plants were regenerated from the protoplast-derived calli on 1/2MS medium containing 2.5–10 g/l sucrose or 5–10 g/l glucose. These results suggest that the kinds of sugar and concentration are important parameters affecting protoplast isolation, proliferation and plant regeneration in L. x formolomgi hort.

Mycophenolic acid production by drug-resistant and methionine or glutamic-acid requiring mutants of Penicillium brevicompactum.

Penicillium brevicompactum ATCC 16024 produced 1.7 g/1 of mycophenolic acid (MPA) in the culture medium. Various drug-resistant mutants, showing resistance to such as polyene antibiotics, chemotherapeutic agents, redox indicator and surfactants, were derived from the fungus. Most of the mutants produced 2.0 ~2.5 g/1 of MPA. A clofibrate and dodecyltrimethylammonium chloride double resistant mutant, No. 4–23–11, produced 4.7 g/1 of MPA. A monofluoroacetic acid resistant strain, No. 5–1, derived from No. 4–23–11 produced 5.3 g/1 of MPA.A methionine auxotroph, M-l, derived from ATCC 16024, produced 4.0 g/1 of MPA. A glutamate auxotroph, G-42, derived from strain No. 4–23–11 produced 5.8 g/1 of MPA. G-42 grew on l-aspartate instead of l-glutamate, and showed one-third the pyruvate carboxylase activity of the parent. Another glutamate auxotroph, G-78, did not produce MPA but accumulated 1.5 g/1 of acetate in the culture medium, and showed one-fifth the citrate synthase activity of the parent strain.

Development of Fertilized Starfish Eggs in Which Cytokinesis is Prevented by Iturin A–2

Iturin A–2, a cyclic peptide obtained from cultures of Bacillus subtilis inhibited cell division but not nuclear divisions of fertilized eggs of the starfish Asterina pectinifera, resulting in the formation of non‐cleaving eggs containing all the embryonic nuclei in a common cytoplasm. Fertilized eggs in which cleavage was prevented by iturin A–2 (50μg/ml) synthesized DNA, RNA and protein at comparable rates to those of normal embryos up to the onset of blastulation. In single‐cell embryos, however, elevation of the level of transcription, which is a characteristic marker of blastulation, did not take place and the chromatin masses eventually dispersed in the cytoplasm.

Relationship between Chemical Structure and Plant Growth-regulating Activity of Amino Acid Related Compounds

Four types of amino acid related compounds were examined on their plant growth-regulating activity. These compounds were N-acylamino acids (N-acyl), N-alkylamino acids (N-alkyl), amino acid higher alkyl esters (ester) and amino acid higher alkyl amides (amide). Every compound, when the number of carbon atoms of the acyl or alkyl radical was 10 to 12, was most effective in inhibiting the root and shoot elongation of rice plant (Oryza sativa L.) in Petri dish. Ester and amide were much more effective than N-acyl and N-alkyl. Ester and amide also showed herbicidal activity against barnyardgrass (Echinochloa crusgalli) grown in pot filled with paddy soil and irrigated, especially, lauryl dl-valinate-HCl being most effective.

Isolation of iturin A-2 as an inhibitor of cytokinesis of starfish embryos

Abstract 1. 1. We have purified substances from the bacterium Bacillus subtilis that inhibit the cell division of embryos of the starfish, Asterina pectinifera , and which are identified as iturin A-2 and closely related cyclic peptides. 2. 2. Iturin A-2 at a concentration of 12.5 μg/ml or greater inhibited cell division of the fertilized starfish egg but not nuclear division, resulting in formation of a multinueleated unicellular embryo. 3. 3. Iturin A-2 microinjected into a fertilized egg was ineffective in halting cell division. 4. 4. From the results of these experiments, we conclude that the arrest of cleavage produced by iturin A-2 is due to the inability of the cell membrane to make a furrow at cytokinesis.